Mechanisms of chloride (Cl–) exclusion in rootstocks of grapevine (Vitis spp.) were studied using a strong Cl–-excluding rootstock (140 Ruggeri) and a poor Cl–-excluding rootstock (K51–40). Xylem Cl– concentration in potted whole grapevines of 140 Ruggeri treated with 50 mM Cl– was 6.8-fold lower than that in K51–40. Five-fold lower total shoot Cl– in salt-treated 140 Ruggeri grapevines relative to that in K51–40 after 50 days, when shoots were of similar biomass, was unrelated to water use. Unidirectional influx of 36Cl– into excised roots (up to 30 min), and uptake of 36Cl– in roots of intact rooted-leaves (up to 180 min) in 10 mM Cl– was similar between the genotypes. However, net accumulation of Cl– by excised roots of K51–40 up to 180 min in 10 mM Cl– was significantly higher than that of 140 Ruggeri. Intact rooted-leaves of 140 Ruggeri in 10 mM Cl– accumulated higher Cl– concentrations in roots, and had a lower percentage of total plant 36Cl– accumulation in the shoot (petiole plus lamina) than those of K51–40. The greater Cl– exclusion capacity of 140 Ruggeri appears to be associated with restricted entry of Cl– to xylem and lower root to shoot Cl– transport.